ECS) volume shrinkage, causes ECS hypertonicity. We hypothesize that ECS hypertonicity will trigger upregulation of Aquaporin 1 (AQP1) in the neurons and pain fibers of Laminae I and II. It has been shown that AQP1 is important for normal pain processing in mouse dorsal horns. We hypothesize that small interfering RNA specific for rat AQP4 (AQP4 siRNA) will block persistent up-regulation of AQP4 in chronically injured spinal cords. Our goal is to test the ability of AQP4 siRNA to alleviate CNP after SCI: (a) by reducing astrocytic swelling, and (b) by decreasing AQP1 expression in pain fibers. We will first establish optimal AQP4 siRNA administered intrathecally for 2 weeks (day 21-35 after SCI) using AQP4 Western blot analyses;and than test the effect of AQP4 siRNA on: (1) CNP (forelimb, trunk and hindlimb mechanical allodynia);(2) astrocytic swelling in chronically injured spinal cords using electron microscopy and by measuring water content;(3) AQP1 expression levels in Laminae I and II, using immunohistochemistry, and (4) test the effect of AQP4 siRNA on astrocytic activation by measuring GFAP levels using Western blots and immunohistochemistry. Our hypothesis on the involvement of AQP4, AQP1 and impaired osmoregulation in the pathogenesis of neuropathic pain conditions after SCI is novel and has not been previously studied. Project Description Page 6